MiR-21 in extracellular vesicles contributes to the growth of fertilized eggs and embryo development in mice

Abstract

Human preimplantation embryo development is susceptible to high rates of early embryo wastage. We determined the miR-21 expression of extracellular vesicles (EVs) in fertilized eggs and embryos of varying stages and their response to miR-21 microinjection. Sexually mature female and male mice were mated. Next, the expression and immunohistochemistry intensity of surface markers (CD9 and CD63) of EVs were detected in pregnant and non-pregnant mice. Exosomes were co-cultured with embryos for detection of blastocyst formation rate, and embryo apoptosis. Moreover, the expressions of Bcl-2 associated X protein (Bax), B cell lymphoma 2 (Bcl-2), and octamer-binding transcription factor-4 (Oct4) were determined. Finally, we detected miR-21 expression in EVs of uterus in pregnant mice, in embryos after embryo implantation and after embryo co-cultured with exosomes in uterine luminal fluid. MiR-21 was up-regulated in EVs of uterus, and higher immunohistochemistry intensity of CD9 and CD63, suggesting more EVs secreted in uterine luminal fluid in pregnant mice. After microinjection, miR-21 inhibitor suppresses embryo development of mice. Moreover, embryos co-cultured with exosomes display higher blastocyst formation rate, reduced apoptotic rate of embryos in pregnant mice. In addition, miR-21 was down-regulated with the development of embryos after embryo implantation, while miR-21 expression in embryos was up-regulated by exosomes in uterine luminal fluid in the pregnant mice. Increased miR-21 expression in EVs of uterus and increased miR-21 expression after implantation, which indicate the key role in the growth of fertilized eggs and embryo development in mice.

Embryotrophic effects of extracellular vesicles derived from outgrowth embryos in pre‐ and peri‐implantation embryonic development in mice

“…In our ongoing study, we found that differentially expressed miRNAs in EVs from outgrowth embryos, comparing those from blastocysts—including miR‐30d, let‐7b, and miR‐429—have functions on various biological processes such as cell communication, cell growth, and proliferation, death, system, and embryo development. Increased expression of miR‐21 was detected in EVs from uterine luminal fluid or pregnant mice have roles in regulating of embryonic development and apoptotic genes expressions such as Bcl2 and Bax identified by loss of function assay (Lv et al, ). Recently, roles of let‐7 family and miR‐429 have been revealed in cell proliferation, apoptosis, implantation, and development process (Al‐Nakhle et al, ; Johnson et al, ; Schickel, Park, Murmann, & Peter, ; Wang et al, ; Zhang et al, ).…”

“…Increased expression of miR-21 was detected in EVs from uterine luminal fluid or pregnant mice have roles in regulating of embryonic development and apoptotic genes expressions such as Bcl2 and Bax identified by loss of function assay(Lv et al, 2018). Recently, roles of let-7 family and miR-429 have been revealed in cell proliferation, apoptosis, implantation, and development process(Al-Nakhle et al, 2010;Johnson et al, 2005;Schickel, Park, Murmann, & Peter, 2010;Wang et al, 2013;Zhang et al, 2015).…”

The Oviductal Extracellular Vesicles’ RNA Cargo Regulates the Bovine Embryonic Transcriptome

“…Overall, the results revealed a network of genes and their regulatory miRNAs that suggested a mode of action of transferred oEV cargo inducing changes in embryonic gene expression which lead to a decrease in apoptosis of embryonic cells and improved embryo viability. For a variety of the identified miRNAs, a positive effect on embryonic development has been shown, for example miR-23a-3p with increased expression in outgrowth embryos in the mouse [75], and miR-21-5p with positive effects on the development of murine embryos [76]. In-vitro-produced bovine embryos cultured in the presence or absence of oEVs for 8 days were used for transcriptomic analysis by RNA sequencing in this study.…”

Small RNA-seq analysis of exosomes from porcine uterine luminal fluid during peri-implantation

“…Among these miRNAs, ssc-miR-21-5p was the most abundant in all libraries. MiR-21 (miR-21-5p) in extracellular vesicles is conducive to embryo development and prevention of high embryo death rate [32]. MiR-21-5p was also found in ovine mammary gland [33] and milk [34] during early pregnancy.…”

“…The microvesicle-like particles are present in saliva, plasma, milk, urine, semen, uterine luminal fluid and other biological fluids [11][12][13][14]. Moreover, these microvesicles have been isolated and characterized from the ULF in human [15], mice [16], cattle [17,18], and sheep [19]. However, porcine ULF exosomes have not been isolated and identified so far.…”